single strand, negative
sense RNA Viruses
Elliot J. Lefkowitz
Contact Information: Elliot Lefkowitz, Ph.D.
Associate Professor, Microbiology
• Email
• ElliotL@uab.edu
• Web Site
• http://www.genome.uab.edu
• Office
• BBRB 277A
• Phone
• 934-1946
Objectives
• To understand the fundamental common and
distinguishing properties of (-) ssRNA viruses
• To understand the basic replication strategies
of (-) ssRNA viruses
• To be able to identify human pathogens that
belong to (-) ssRNA virus families, and some
of their biological and pathogenic properties
Reading
• Medical Microbiology, Murray et al. 6th
Edition
• General classification
• Chapter 4
• RNA virus properties and replication
• Chapters 58, 59, 60, 63
• Pathogenesis
• Chapters 48, 67
Slide References
• Fields Virology, 5th Edition
• Viruses and Human Disease
• Strauss and Strauss
• University of Leicester - Virology Online
• http://wwwmicro.msb.le.ac.uk/3035/index.html
• International Committee on Taxonomy of
Viruses
• The 9th ICTV Report
• Primary literature
Virus classification
The Virus World
The (-) RNA Virus World
RNA Virus Genome Structure
• Number of strands
• Single or double stranded
• Strand polarity
• Positive, negative, or ambisense (both + and -)
• Positive (Plus) sense denotes the coding
(mRNA) strand
• Number of segments
• Single or multi-segmented
single strand RNA virus genome
polarity
(+) sense RNA virus
virion RNA (+) sense
virus mRNA (+)
sense
5’
3’
translation
(-) sense RNA virus
virion RNA (-) sense
virus mRNA (+) sense
3’
5’
transcription
translation
5’
3’
Negative/Ambisense ssRNA
Viruses
Properties of (-) sense ssRNA Viruses
• Enveloped virion
• Helical nucleocapsid
• Negative-sense, linear, single segment RNA genome
• Bornaviruses, Filoviruses, Rhabdoviruses,
Paramyxoviruses
• Negative and Ambisense, linear, multi segment RNA
genomes
• Arenaviruses, Bunyaviruses, Orthomyxoviruses
• Cytoplasmic replication
• Exception: Bornaviruses, Orthomyxoviruses
• Genomes are non-infectious
• An initial round of transcription is required for genome
replication
• Virion must contain proteins required for transcription
Order: Mononegavirales:
Single segment, (-) sense, ssRNA
•
•
•
•
Bornaviridae
• Bornavirus
Filoviridae
• Marburg virus
• Ebola virus
Paramyxoviridae
• Paramyxovirinae
• Henipavirus
• Morbillivirus
• Respirovirus
• Rubulavirus
• Pneumovirinae
• Pneumovirus
• Metapneumovirus
Rhabdoviridae
• Vesiculovirus
• Lyssavirus
Multi-Segment, (-) sense ssRNA
viruses
• Orthomyxoviridae
• Influenzavirus A
• 8 genome segments
• Influenzavirus B
• 8 genome segments
• Influenzavirus C
• 7 genome segments
• Isavirus
• 8 genome segments
• Thogotavirus
• 6 genome segments
Multi-Segment, Negative and
Ambisense ssRNA viruses
• Arenaviridae
• Two ambisense RNA segments
• Bunyaviridae
• Three RNA segments
• Both negative-sense and ambisense
segments
• Depends on genus
The Virus
Virion, Genome, Proteins
Viral Proteins
• Attachment/entry
• G – Membrane glycoprotein
• F – Fusion protein
• H – Hemagglutinin
• N – Neuraminidase
• Structural/Assembly
• M – Matrix
• Underlies lipid bylayer
• Replication
• N – nucleocapsid protein
• P – Phosphoprotein
• L – RNA dependent RNA polymerase
Rhabdovirus Virion
Virus replication Machinery
• Proteins
• RNA-dependent RNA-polymerase (RdRp)
• Transcription
• Replication
• Nucleocapsidprotein (N)
• Encapsidates RNA
• Forms helical nucleocapsid
• P protein
• Phosphoprotein - polymerase cofactor
• Forms complexes with N and L
• Binds to RNA termini
• RNA Genome
Genome Organization Mononegavirales
Filoviridae
Paramyxoviridae
Rhabdoviridae
Genome Organization
Arenaviridae
Bunyaviridae
Influenza A Genome Structure
Virus Coding Strategies
• Individual ORFs
• Multiple transcripts with transcription
attenuation
• Polyprotein processing
• Single transcript to Large polyprotein:
Proteolytic processing
• RNA Editing
• Insertion/deletion of additional residues (at
a specified site) altering the reading frame
• Multiple ribosomal initiation sites
• Stop codon read-through
Virus Replication
RNA-dependent RNA Polymerase
(RdRp – L Protein)
• Catalytic subunit of the polymerase complex
• Polymerization of nucleotides
• Transcription of mRNA
• Capping
• Methylation
• Polyadenylation
• Genome Replication
• Most conserved protein between the
mononegavirales virus families
Source of the RNA-dependent RNA
Polymerase
• Host cells do not have a suitable one
• Therefore the virus must provide its own
• RNA viruses use 2 different strategies to provide
the RdRp:
• Synthesized immediately upon entry and
unpackaging of the virion into the cell
(positive-sense viruses)
• Therefore protein synthesis is the first
step in the replication process
• Packaged within the virion (negative-sense
viruses)
• Therefore mRNA transcription is the first
step in the replication process
VSV Transcription & Replication
(-) sense ssRNA virus
Human Pathogens
Major Viral Target Tissues
Arenaviruses/Bunyaviruses
Arenavirus and Bunyavirus Disease
• Arenaviruses
• Mostly rodent viruses
• Human zoonoses
• Junin virus
• Argentine hemorrhagic fever
• Lassa Fever
• Bunyaviruses
• Large group of arthropod-borne viruses
• Human pathogens – hemorrhagic fever
• Hantaviruses
• Rodent-borne
• Pulmonary Syndrome/Hemorrhagic fever
• Rift Valley Fever virus
• Mosquito-borne virus
Filoviruses
Filovirus Disease
Rhabdoviruses
Rhabdoviruses
Rabies virus Pathogenesis
Paramyxoviruses
Paramyxoviruses
Human Respiratory Syncytial virus
• Major cause of lower respiratory tract infections
• Rarely life-threatening
• Individuals get repeat infections
• Highly infectious
• Spread is by exchange of respiratory
•
•
secretions
• Infection confined to respiratory tract
Globally: 100,000,000 infections/year
• 200,000 deaths/year
In USA: All infants by age of 4 years are infected
• 100,000 hospitalizations/year
• Estimated cost of $300,000,000/year (1985)
• 25-50% of hospital staff infected during
outbreaks
Measles virus
• Extremely infectious
• Spreads through contact with respiratory
secretions
• Victims are infectious before symptoms are
evident
• Develops systemic infection
• Globally: 45,000,000 infections/year
• 1,000,000 deaths/year
• In USA: Infections are rare
• Occasional epidemic in unvaccinated populations
• MMR (Measles, mumps, and rubella) vaccine
highly effective (2 shots)
Neurologic Complications of
Measles
Acute Disseminated Encephalomyelitis
Measles Inclusion Body Encephalitis
Subacute Sclerosing Panencephalitis
Orthomyxoviruses
Orthomyxoviruses
• Influenza
• A: Mild to severe disease involving upper and
•
•
especially lower respiratory tract
• B: Similar spectrum of illness to A but
generally more mild
• C: Sporadic upper respiratory illness in
humans
• 96% of human adults have antibodies
Thogotovirus
• Natural host: Ticks
• Also infects: Humans, cattle, goats, waterfowl,
etc.
Isavirus
• Infectious salmon anemia virus
Schematic diagram of influenza A
viruses
G Neumann et al. Nature 000, 1-9 (2009) doi:10.1038/nature08157
M2 Ion Channel
• Involved in virion
uncoating
• Highly conserved
• Target for amantadine
Hemagglutinin
Neuraminidase
• Virion release from cell
membrane
• Cleavage of sialic
acid from cell
membrane thus
preventing binding
by HA
• Target for Oseltamavir
(Tamavir) and
Zanamivir (Relenza)
Variation and evolution
Influenza virus
Influenza Virus Variation
• Antigenic drift
• Amino acid changes
• Antigenic shift
• Reassortment/exchange of genome
segments between strains
• Recombination
• Detected but rare
Reassortment
Genesis of swine-origin H1N1 influenza
viruses
G Neumann et al. Nature 000, 1-9 (2009) doi:10.1038/nature08157
Why Pigs?
• Susceptible to infection by influenza virus
• Express both human- and avian-like
influenza virus receptors on their tracheal
epithelial cells
• Swine may therefore be acting as a “mixing
vessel” for the production, replication, and
transmission of novel influenza virus
reassortments
US Influenza Surveillance 20042008
US Influenza Surveillance 20082009
US Influenza Surveillance 20102011
Fighting back
• Antiviral drugs
• Neuraminidase inhibitors
• Oseltamavir (Tamavir) and Zanamivir
•
(Relenza)
• Active against influenza A and B
• Ion channel blockers
• Amantidine and rimantidine
• Prevent release and subsequent transport
of the virus RNP
• Active only against Influenza A
Vaccines
• Inactivated
• Live attenuated
Antiviral Resistance
Antiviral Resistance 2008 - 2009
Antiviral Resistance 2010 - 2011
Vaccine Development
• Inactivated vaccine (TIV)
• Produced from seed stocks in eggs
• Live-attenuated vaccine (LAIV)
• Administered as a nasal spray
• Vaccines contain three viruses
• H3N2; H1N1; B
• Exact strains used change each year
• Strain choice determined by data collected by
WHO on currently circulating strains
• Decision on composition made in February
and September
Vaccine Strains for the
2009-2010 and 2010-2011 Seasons
• 2009 – 2010 Seasonal Vaccine
• A components unchanged from 2008-2009
• B component changed toB/Brisbane/60/2008
• Related to B/Victoria
• 2009 Supplementary Vaccine
• A/California/7/2009 (H1N1)
• 2009 pandemic influenza A (H1N1) virus
• 2010 – 2011 AND 2011 – 2012 influenza A (H1N1)
virus
• A/California/7/2009 (H1N1)-like virus (99.8% of
2010-2011 viruses match)
• A/Perth/16/2009 (H3N2)-like (96.8% of 2010-2011
viruses match)
• B/Brisbane/60/2008 (94% of 2010-2011 viruses
match)
And finally, how is influenza
spread between humans
and pigs?